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Impaired learning and memory are common in epilepsy syndromes of childhood. Clinical investigations suggest that the developing brain may be particularly vulnerable to the effects of intractable seizure disorders. Magnetic resonance imaging (MRI) studies have demonstrated reduced volumes in brain regions involved in learning and memory. The earlier the onset of an epilepsy the larger the effects seem to be on both brain anatomy and cognition. Thus, childhood epilepsy has been proposed to interfere in some unknown way with brain development. Experiments reported here explore these ideas by examining the effects of seizures in infant mice on learning and memory and on the growth of CA1 hippocampal pyramidal cell dendrites. Fifteen brief seizures were induced by flurothyl between postnatal days 7 and 11 in mice that express green fluorescent protein (GFP) in hippocampal pyramidal cells. One to 44 days later, dendritic arbors were reconstructed to measure growth. Spatial learning and memory were also assessed in a water maze. Our results show that recurrent seizures produced marked deficits in learning and memory. Seizures also dramatically slowed the growth of basilar dendrites while neurons in litter-mate control mice continued to add new dendritic branches and lengthen existing branches. When experiments were performed in older mice, seizures had no measureable effects on either dendrite arbor complexity or spatial learning and memory. Our results suggest that the recurring seizures of intractable childhood epilepsy contribute to associated learning and memory deficits by suppressing dendrite growth.

In the primary auditory cortex (AI), a pure tone evokes propagating activity along a strip of the cortex. We have previously shown that focal activation of AI triggers autonomously propagating activity that resembles tone-evoked activity (Song et al., 2006). Because a focal spontaneous activity is expected to trigger similar activity propagation, spontaneous activity resembling tone-evoked activity may exist in AI. Here we tested this possibility by optical imaging of AI in guinea pigs. After obtaining tone-evoked activities, we made long-duration optical recordings (9-40s) and isolated spontaneous activities from respiration and heartbeat noises using independent component analyses. Spontaneous activities were found all over AI, in all animals examined. Of all spontaneous events, 33.6% showed significant correlation in spatio-temporal pattern with tone-evoked activities. Simulation using a model that captures the temporal feature of spontaneous response in single channels but sets no constraint among channels, generated no spontaneous events that resembled tone-evoked activations. These results show the existence of spontaneous events similar in spatio-temporal pattern to tone-evoked activations in AI. Such spontaneous events are likely a manifestation of cortical structures that govern the pattern of distributed activation in AI.

X-linked dystonia-parkinsonism (XDP) is a movement disorder endemic to the Philippines. The disease locus, DYT3, has been mapped to Xq13.1. In a search for the causative gene, we performed genomic sequencing analysis, followed by expression analysis of XDP brain tissues. We found a disease-specific SVA (short interspersed nuclear element, variable number of tandem repeats, and Alu composite) retrotransposon insertion in an intron of the TATA-binding protein-associated factor 1 gene (TAF1), which encodes the largest component of the TFIID complex, and significantly decreased expression levels of TAF1 and the dopamine receptor D2 gene (DRD2) in the caudate nucleus. We also identified an abnormal pattern of DNA methylation in the retrotransposon in the genome from the patient’s caudate, which could account for decreased expression of TAF1. Our findings suggest that the reduced neuron-specific expression of the TAF1 gene is associated with XDP.

We tested the possibility of using a high-power monochromatic InGaN light-emitting diode (LED) as an excitation light source for real-time optical imaging using the voltage-sensitive dye RH-795. Driven with a custom-designed, non-feedback-controlled constant-current circuit, the LED generated stable light with rapid on/off. The LED was comparable with commonly used halogen lamps in exciting RH-795. Acoustically evoked responses in the auditory cortex recorded with the two light sources were highly similar. Our results thus suggest that a high-power LED can be successfully used as an excitation light source for voltage-sensitive dyes, without the need of optical filters and shutters.

Circadian clock genes directly regulate expression of the Na + /H + exchanger NHE3 in the kidney. Background Daily rhythms in mammalian physiology are generated by a transcription/translation feedback loop orchestrated by a set of clock genes. However, little is known about the molecular cascade from the clock gene oscillators to cellular function. Methods The mRNA expression profiles of NHE3 and clock genes were examined in mice and rat kidneys. First, luciferase assays followed by a site directed mutagenesis of an E-box sequence were used to assess the CLOCK:BMAL1-transactivated NHE3 promoter activity. A direct binding of CLOCK:BMAL1 heterodimers to an E-box sequences of NHE3 promoter was confirmed by electrophoretic mobility shift assay (EMSA). Results We present evidence that renal tubular NHE3 , the Na + /H + exchanger critical for systemic electrolyte and acid-base homeostasis, is a clock-controlled gene regulated directly by CLOCK:BMAL1 heterodimers in kidneys. NHE3 mRNA level in rat kidney displayed circadian kinetics, and this circadian expression was severely blunted in homozygous CRY1/2 double-deficient mice, suggesting that the transcriptional machinery of peripheral clocks in renal tubular cells directly regulates the circadian expression of NHE3 . By analyzing the 5′ upstream region of the NHE3 gene, we found an E box critical for the transcription of NHE3 via the CLOCK:BMAL1-driven circadian oscillator. The circadian expression of NHE3 mRNA was reflected by oscillating protein levels in the proximal tubules of the rat kidney. Conclusion NHE3 should represent an output gene of the peripheral oscillators in kidney, which is regulated directly by CLOCK:BMAL1 heterodimers.

Human T-cell leukemia virus type I (HTLV-I) provirus load differs more than 100-fold among carriers and a high provirus load in the peripheral blood mononuclear cells (PBMCs) is regarded as a risk factor for both preleukemic states and inflammatory diseases including HTLV-I-associated myelopathy (HAM). We examined polymorphisms in the genes for tumor necrosis factor (TNF), TNF receptor type 1 and 2, lymphotoxin (LT)-alpha, interleukin (IL)-1beta, IL-6, IL-10, monocyte chemoattractant protein (MCP)-1, and mannose binding protein (ManBP) in 143 HTLV-I carriers whether these polymorphisms affect the provirus load in the PBMCs of carriers. No significant association was observed between these polymorphisms and the provirus load. Homozygotes for a ManBP-variant allele, however, showed a tendency for the decreased number of provirus load. When combined, the data on the alleles of LT-alpha and MCP-1, HTLV-I carriers having high producer alleles of both genes showed a trend for increased provirus load. These data suggest that inflammation or an active immune response may induce an increased amount of HTLV-I-infected T cells, leading to a high provirus load.

We studied cytokine gene polymorphisms in the promoter region, including interleukin (IL)-6, IL-1beta, and IL-10, in Japanese patients with human T-cell leukemia virus type I (HTLV-I) associated myelopathy (HAM) (n = 65), asymptomatic HTLV-I carriers (n = 143), and HTLV-I seronegative, normal controls (n = 160). There was a significant difference between HAM patients and HTLV-I carriers in the distribution of IL-6 promoter polymorphism at position -634 (chi(2) = 9.90, p = 0.0071). The IL-6 genotype was also significantly different between HAM patients and normal controls (chi(2) = 11.53, p = 0.0033), while a similar distribution was observed in IL-1beta and IL-10 polymorphisms among HAM patients, carriers, and normal controls. The results suggest that IL-6 gene region may contribute to susceptibility to HAM, and that aberrant cytokine productions could be involved in the development of HAM.

We investigated the effects of selegiline and desmethylselegiline on synthesis of neurotrophic factors in cultured mouse astrocytes. Treatment with 2 mM selegiline for 24 h increased the contents of NGF, BDNF, and GDNF in the culture medium 26-, 1.7-, and 4.2-fold over the control, respectively. With this drug the maximum relative mRNA levels of NGF, BDNF, and GDNF were 6.2-fold at 2 h, 3.4-fold at 6 h, and 2.7-fold at 2 h, respectively. Selegiline at 0.2 mM completely inhibited the MAO activity, but had no effect on the content of neurotrophic factors, suggesting that stimulation of neurotrophic factors by selegiline is independent of MAO-B inhibition. Desmethylselegiline at 1.68 mM for 24 h elevated the NGF, BDNF, and GDNF contents 4.1-, 1.7-, and 2.4-fold over the control, respectively; and the relative transcript levels of NGF, BDNF, and GDNF reached 2.6-fold at 2 h, 1.7-fold at 6 h, and 1.8-fold at 2 h, respectively. These findings suggest that selegiline and desmethylselegiline may protect neurons by up-regulating endogenous NGF, BDNF, and GDNF synthesis.

Apomorphine, a D1/D2 dopamine agonist, is an anti-parkinsonian drug. We examined the effects of apomorphine on synthesis of neurotrophic factors in cultured mouse astrocytes. After 24 h incubation with apomorphine, NGF and GDNF contents in the culture medium increased to 122-fold and 1.8-fold of the control, respectively; whereas the BDNF content did not change significantly. In Northern blot analysis, expression of NGF mRNA in astrocytes reached the maximum level at 6 h after addition of the drug. By semiquantitative RT-PCR analysis, the GDNF transcript level was found to reach 2.9-fold of the control level at 15 h. These results suggest that apomorphine may exert neuroprotective effects by stimulation of NGF and GDNF synthesis in astrocytes.

We studied genetic polymorphisms in the promoter region (position -511) and exon 5 (position +3953) of the interleukin (IL)-1beta gene in 122 Japanese patients with Parkinson's disease (PD) and 112 controls. We also examined polymorphisms in the IL-1alpha and the IL-1 receptor antagonist genes. No significant difference was found in these genetic markers between PD patients and controls. However, PD patients with homozygotes for allele 1 at position -511 of the IL-1beta gene (IL-1B-511*1), a low producer of IL-1beta, were significantly earlier in the disease onset than those with the IL-1B-511*2 homozygotes, a high producer of IL-1beta. This suggests that IL-1beta might play a role, possibly a protective effect for dopaminergic neurons, in PD. Further population and functional studies are necessary to clarify the role of IL-1beta in PD patients.

We examined the circadian rhythm of urinary sodium excretion and the effects of sodium restriction on rhythm in both dipper and nondipper types of essential hypertension. Patients (n = 26) with essential hypertension were maintained on relatively high- (10 to 12 g/d of sodium chloride) and low-sodium (1 to 3 g/d) diets for 1 week each. On the last day of each diet, 24-hour blood pressures (BPs) were measured every 30 minutes noninvasively with an automatic device, and on the last 3 days, urinary samples were collected for both daytime (7:00 AM to 9:30 PM) and nighttime (9:30 PM to 7:00 AM). Eight patients were classified as dippers based on a more than 10% reduction in mean arterial pressure (MAP) from daytime to nighttime on a high-sodium diet, and 18 patients were classified as nondippers. A nocturnal decrease in urinary sodium excretion rate (U(Na)V) on the high-sodium diet was observed in dippers (from 7.5 +/- 2.1 during the day to 5.3 +/- 2.5 mmol/h at night; P < 0.0001), but not in nondippers (6.7 +/- 2.1 v 7.6 +/- 2.3 mmol/h; not significant). In nondippers, the night-day ratio of sodium excretion was significantly reduced from 1.2 +/- 0.4 to 0.8 +/- 0.3 (P < 0.003) by sodium restriction; at the same time, the night-day ratio of MAP was reduced from 0.98 +/- 0.04 to 0.93 +/- 0.05 (P < 0.05). In dippers, the night-day ratios of MAP and U(Na)V were not affected by sodium restriction, and both ratios remained constant at less than 1. Before sodium restriction, the night-day ratio of sodium excretion correlated with that of MAP (r = 0.78; P < 0.0001), whereas there was no significant correlation (r = -0.05) after sodium restriction. These findings showed that the circadian rhythm of renal sodium excretion differed between the two types of essential hypertension. The enhanced nocturnal natriuresis and diminished nocturnal BP fall on a high-sodium diet, recognized in nondippers, were both normalized by sodium restriction, resulting in circadian rhythms with nocturnal dips in U(Na)V and BP.

We studied genetic polymorphisms in the tumor necrosis factor (TNF) region as well as HLA-DRB1 of 42 patients with Western-type multiple sclerosis (MS) and 38 with Asian-type MS in Japan. The sex ratio (Female:Male) was significantly higher in Asian than in Western type MS (3.8 vs. 1.3, P =0.038). The frequency of HLA-DRB1*1501 allele in the Western-type MS group increased significantly compared with the control group, while Asian-type MS and control groups showed similar distribution in the frequencies of HLA-DRB1 alleles. No significant differences existed in the TNF region, however, including TNF-a microsatellite alleles. The results suggest that MS in Asians may present two different clinical and immunogenetic manifestations.

Campylobacter jejuni is a major pathogen preceding Guillain-Barré syndrome (GBS), and most C. jejuni isolates from GBS patients belong to Penner serotype 19 (heat-stable; HS-19). We analyzed sixteen independent clinical isolates from GBS patients, twelve of which belonged to HS-19, three to HS-2, and one to HS-4, using PCR-based RFLP analysis of a flagellin-A (flaA) gene. Two isolates from patients with Miller Fisher syndrome (MFS), and 27 from patients with uncomplicated enteritis were also examined. All HS-19 isolates, regardless of GBS, showed an identical pattern (Cj-1) by RFLP typing and were distinguishable from those of the other Penner serogroups. In contrast, HS-2 and HS-4 isolates were divided into several different RFLP groups, suggesting HS-19 strains are genetically distinctive among C. jejuni isolates. A DNA fingerprinting method also failed to detect any specific band pattern for GBS-related C. jejuni isolates. We examined relationships among anti-GM1 antibody titres in the sera of GBS patients, clinical forms of GBS, serotype of C. jejuni, and the presence of GM1-like structures in lipopolysaccharide (LPS) components from C. jejuni isolates by immunoblotting. HS-19 related GBS was significantly associated with elevated anti-GM1 antibody titers in the sera of the patients, but not associated with any clinical pattern of GBS. No significant correlations were found between anti-GM1 antibody and the pattern of disease, or between GBS-related C. jejuni strains and the presence of GM1-like structures. HS-19 strains seem to be unique among C. jejuni isolates, and HS-19-related GBS may provide an excellent model for clarification of the pathogenesis of GBS.

T cell lines were established from the cerebrospinal fluid (CSF) lymphocytes of three patients with human T lymphotropic virus type I (HTLV-I)-associated myelopathy (HAM). To elucidate the possible changes of the provirus nucleotide sequences integrated in HAM-derived T cell line cells, DNA from these cell was digested with Pst I, which cleaved the provirus genome of HTLV-I at several sites, and three common bands were detected by Southern blot analysis in all cases. These bands were identical in size to those detected in T cell lines established from peripheral blood lymphocytes of adult T cell leukemia (ATL) patients. These results were confirmed with restriction enzymes Hin III and Bam HI. These findings suggest that the provirus genome detected in T cell lines derived from CSF of HAM patients is identical to HTLV-I.

Progressive subcortical vascular encephalopathy (PSVE) usually occurs in elderly individuals, suffering from hypertension. We here describe a male, born of consanguineous parents, who first showed signs of PSVE at the age of 30. Despite the absence of hypertension or known metabolic causes, the degenerative cerebral vascular disease developed progressively. Several cases, surprisingly identical to the one reported here, were traced using Japanese medical records. They are clinically characterized by: early onset of PSVE (at age 25-30), absence of persistent hypertension, diffuse alopecia since youth, spondylitis deformans with early onset, often so severe as to necessitate surgery, and the possible existence of an autosomal recessive transmission. Cases with these features appear to constitute a distinct clinical entity, possibly a new form of premature aging syndrome.